Evidence has been presented suggesting that hemoglobin (Hb) may occupy a functional role as an oxygenase enzyme. It has been reported that Hb is capable of catalyzing the oxidation of aniline to p-aminophenol in a reconstituted enzyme system consisting of Hb, NADPH-cytochrome P-450 reductase, NADPH and molecular 02, as well as in lysed and intact red cells and whole human blood. Hb02 was required for catalytic activity and binding of the substrate to Hb and catalytic reactivity have been correlated with substrate-induced alterations of Hb affinity for 02. The scope of such Hb-catalyzed oxidation reactions has been reported to include N-demethylase and 0-dealkylase activity. Fetal erythrocytes are 3-5 times more active in the conversion of aniline to p-aminophenol than normal adult erythrocytes. A variety of volatile organic solvents and other organic materials with significant vapor pressure are commonly present in various industrial and research environments. Chronic or acute exposure to these agents is followed by rapid absorption into the circulatory system and may result in altered Hb02 affinity while converting Hb into and 02-activating system (hemeoxygenase) capable of catalyzing the formation of either innocuous metabolites or toxic products such as reactive intermediates (epoxides, hydroxylamines) or reactive 02 species (superoxide anion, H202) which may compromise the functioning and integrity of the red cell. Little information however, is available on the effects of aromatic hydrocarbons on Hb02 affinity, the degree or extent of Hb-catalyzed oxidation reactions, or the subsequent effects of such interactions and/or oxidative processes on red cell integrity or functioning. In addition, virtually no information exists concerning the propensity of Hb in red cells to catalyze the formation of reactive or toxic metabolites such as epoxides or hydroxylamines. Thus, the overall objective of this proposal includes complete characterization of the effects of these agents on Hb as an isolated hemeprotein, in a reconstituted enzyme system, and in lysed and intact red cells in order to determine the nature, scope and extent of aromatic hydrocarbon-induced alterations in Hb02 affinity, Hb-catalyzed oxidation reactions, reactive metabolite formation, and the subsequent effects of hemeoxygenase activity and related product formation on red cell function and in integrity.